The gene encoding Runx1 (AML1) is disrupted by the t(8;21) that is associated with ~12% of acute myeloid leukemia in humans. The t(8;21) results in production of a fusion protein containing the N-terminus of Runx1, including the Runt domain, fused to almost all of ETO. The AML1-ETO fusion protein acts as a dominant repressor of core binding factor function, dysregulating the expression of multiple genes required for normal hematopoiesis and, in cooperation with secondary mutations, leads to the development of leukemia. Most t(8;21) patients treated with current therapies who achieve hematological and cytogenetic long-term remission retain AML1-ETO transcripts in their bone marrow which are produced from either leukemic or preleukemic cells that were not eradicated. Thus, although 90% of patients achieve complete remission, 35-40% of these patients relapse within five years due to the resurgence of residual t(8;21) expressing cells. We hypothesize that direct therapeutic targeting of the AML1-ETO protein may reduce the rate of relapse and improve the long- term survival of these patients. We plan to specifically inhibit the activity of the AML1-ETO protein with small molecules. To this end, we have shown that the AML1-ETO:CBF2 interaction is essential for AML1-ETO's leukemogenic and pre- leukemogenic activity. We have begun to develop small molecule inhibitors of the AML1-ETO:CBF2 interaction using structure-based and high throughput screens and well-validated assays. We are employing medicinal chemistry to optimize these leads. We also plan to develop homodimeric and heterodimeric inhibitors from the small molecules that will have selectivity for AML1-ETO versus Runx1 (AML1). All inhibitors will be tested for their activity on mouse and leukemia cell lines, on normal bone marrow cells, and in mouse models for t(8;21) leukemia. PUBLIC HEALTH RELEVANCE: We are proposing to develop small molecule inhibitors of the fusion protein AML1-ETO which is a causative agent in one type of acute myeloid leukemia (AML). Such targeted inhibitors have the potential to improve the long term survival of patients with this leukemia by eliminating the population of leukemia cells which cause relapse to occur. These inhibitors also have the potential to be less toxic than standard chemotherapy, resulting in reduced side effects from treatment.